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Rerouting Cellular Electron Flux To Increase the Rate of Biological Methane Production.

Catlett JL, Ortiz AM, Buan NR - Appl. Environ. Microbiol. (2015)

Bottom Line: In Methanosarcina acetivorans, HdrABC expression caused an increased rate of methanogenesis and a decrease in metabolic efficiency on methylotrophic substrates.When acetate was the sole carbon and energy source, neither deletion nor overexpression of HdrABC had an effect on growth or methane production rates.These results suggest that in cells grown on methylated substrates, the cell compensates for energy losses due to expression of HdrABC with an increased rate of substrate turnover and that HdrABC lacks the appropriate electron donor in acetate-grown cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry, Redox Biology Center, University of Nebraska-Lincoln, Lincoln, Nebraska, USA.

No MeSH data available.


Related in: MedlinePlus

HdrABC uncouples methanogenesis from cell growth. (A) In cell suspension assays, the rate of methane production is dependent on the amount of HdrABC enzyme activity in the cell. (B) The growth rate and methane production rate are correlated by a second-order relationship. (C) Metabolic efficiency of each strain. The values are normalized to the ΔhdrABC mutant strain. (D) HdrABC enzyme activity has a strong negative correlation with metabolic efficiency and fits a direct-reduction mechanism model (R2 = 0.9981). The error bars indicate the standard deviations and may not be visible behind the symbols.
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Figure 4: HdrABC uncouples methanogenesis from cell growth. (A) In cell suspension assays, the rate of methane production is dependent on the amount of HdrABC enzyme activity in the cell. (B) The growth rate and methane production rate are correlated by a second-order relationship. (C) Metabolic efficiency of each strain. The values are normalized to the ΔhdrABC mutant strain. (D) HdrABC enzyme activity has a strong negative correlation with metabolic efficiency and fits a direct-reduction mechanism model (R2 = 0.9981). The error bars indicate the standard deviations and may not be visible behind the symbols.

Mentions: After determining that the pJC1 plasmid can complement the ΔhdrABC mutant growth phenotype, we tested whether overexpression of HdrABC in the parent strain (att::hdrABC*) would have the opposite effect of a ΔhdrABC mutation and result in increased growth and methane production. Decreased expression of HdrABC (by deleting the hdrABC operon) resulted in a 50% decrease in methane production and a 20% decrease in the growth rate. However, instead of increasing growth and methane production, the att::hdrABC* mutant strain we created had a growth rate identical to that of the parent strain (Fig. 2A). When cells were switched from TMA to methanol, the parent had a doubling time of 10.1 ± 0.16 h versus 10.4 ± 0.21 for the att::hdrABC* mutant strain (see Table S3 in the supplemental material). When cells were preadapted to methanol for 25 generations, both the parent and the att::hdrABC* mutant had cell doubling times of 8.5 h (Table 1). When methane production was measured in resting cell suspensions, the att::hdrABC* mutant exhibited a 30% increase in the rate of methane production from methanol versus the parental strain (Fig. 3B). The methanol consumption rate was consistent with the methane production rates. The ΔhdrABC deletion mutant had a 15% decrease in the methanol consumption rate versus the parent strain, the ΔhdrABC att::hdrABC* complemented strain had a rate 16% higher than that of the parent strain, and the att::hdrABC* overexpression strain had a 16% increase in rate compared to the parent. The Hdr enzyme assays, growth phenotypes, and methane rate assays suggest that an increased HdrABC enzyme activity of 12% results in a 30% increase in conversion of methanol to methane without affecting the kinetics of cell growth. Our data show that the rate of methane production is dependent on the rate of Hdr enzyme activity by a direct linear correlation (Fig. 4A). As Hdr enzyme activity (kHdrABC) in the cell is increased, the rate of methane production (kCH4) increases according to equation 1: (1)kCH4=41.817×kHdrABC−295.65


Rerouting Cellular Electron Flux To Increase the Rate of Biological Methane Production.

Catlett JL, Ortiz AM, Buan NR - Appl. Environ. Microbiol. (2015)

HdrABC uncouples methanogenesis from cell growth. (A) In cell suspension assays, the rate of methane production is dependent on the amount of HdrABC enzyme activity in the cell. (B) The growth rate and methane production rate are correlated by a second-order relationship. (C) Metabolic efficiency of each strain. The values are normalized to the ΔhdrABC mutant strain. (D) HdrABC enzyme activity has a strong negative correlation with metabolic efficiency and fits a direct-reduction mechanism model (R2 = 0.9981). The error bars indicate the standard deviations and may not be visible behind the symbols.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4561719&req=5

Figure 4: HdrABC uncouples methanogenesis from cell growth. (A) In cell suspension assays, the rate of methane production is dependent on the amount of HdrABC enzyme activity in the cell. (B) The growth rate and methane production rate are correlated by a second-order relationship. (C) Metabolic efficiency of each strain. The values are normalized to the ΔhdrABC mutant strain. (D) HdrABC enzyme activity has a strong negative correlation with metabolic efficiency and fits a direct-reduction mechanism model (R2 = 0.9981). The error bars indicate the standard deviations and may not be visible behind the symbols.
Mentions: After determining that the pJC1 plasmid can complement the ΔhdrABC mutant growth phenotype, we tested whether overexpression of HdrABC in the parent strain (att::hdrABC*) would have the opposite effect of a ΔhdrABC mutation and result in increased growth and methane production. Decreased expression of HdrABC (by deleting the hdrABC operon) resulted in a 50% decrease in methane production and a 20% decrease in the growth rate. However, instead of increasing growth and methane production, the att::hdrABC* mutant strain we created had a growth rate identical to that of the parent strain (Fig. 2A). When cells were switched from TMA to methanol, the parent had a doubling time of 10.1 ± 0.16 h versus 10.4 ± 0.21 for the att::hdrABC* mutant strain (see Table S3 in the supplemental material). When cells were preadapted to methanol for 25 generations, both the parent and the att::hdrABC* mutant had cell doubling times of 8.5 h (Table 1). When methane production was measured in resting cell suspensions, the att::hdrABC* mutant exhibited a 30% increase in the rate of methane production from methanol versus the parental strain (Fig. 3B). The methanol consumption rate was consistent with the methane production rates. The ΔhdrABC deletion mutant had a 15% decrease in the methanol consumption rate versus the parent strain, the ΔhdrABC att::hdrABC* complemented strain had a rate 16% higher than that of the parent strain, and the att::hdrABC* overexpression strain had a 16% increase in rate compared to the parent. The Hdr enzyme assays, growth phenotypes, and methane rate assays suggest that an increased HdrABC enzyme activity of 12% results in a 30% increase in conversion of methanol to methane without affecting the kinetics of cell growth. Our data show that the rate of methane production is dependent on the rate of Hdr enzyme activity by a direct linear correlation (Fig. 4A). As Hdr enzyme activity (kHdrABC) in the cell is increased, the rate of methane production (kCH4) increases according to equation 1: (1)kCH4=41.817×kHdrABC−295.65

Bottom Line: In Methanosarcina acetivorans, HdrABC expression caused an increased rate of methanogenesis and a decrease in metabolic efficiency on methylotrophic substrates.When acetate was the sole carbon and energy source, neither deletion nor overexpression of HdrABC had an effect on growth or methane production rates.These results suggest that in cells grown on methylated substrates, the cell compensates for energy losses due to expression of HdrABC with an increased rate of substrate turnover and that HdrABC lacks the appropriate electron donor in acetate-grown cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry, Redox Biology Center, University of Nebraska-Lincoln, Lincoln, Nebraska, USA.

No MeSH data available.


Related in: MedlinePlus